Radiation mitigator and method of use thereof

ABSTRACT

Provided are methods useful for preventing and mitigating radiation injury, including acute radiation syndrome, comprising administering to a subject a therapeutically effective amount of a compound represented by formula Ior a pharmaceutically acceptable salt thereof, wherein Z is —O— or —N(H)—.

CROSS REFERENCE TO RELATED APPLICATION

This application claims benefit of priority to U.S. ProvisionalApplication No. 62/316,121, filed Mar. 31, 2016.

BACKGROUND

Ionizing radiation, i.e., radiation with energy high enough to have thepotential of ionizing molecules in a living body, may cause seriousdamage and injury to the cells and tissues of living beings. Ionizingradiation damages tissue by direct ionization, which disrupts moleculesdirectly, and also by producing highly reactive free radicals, whichattack nearby cells. The net effect is that biological molecules sufferlocal disruption; this may exceed the body's capacity to repair thedamage and may cause mutations in cells currently undergoing replicationthat lead to diseases. In consequence, dysfunction of many importantorgans, and even multiple organ failure, can occur, which in turn caneventually lead to death (radiation-induced lethality).

Damaging and harmful effects of radiation can be observed both in thecase of acute high dose exposure and in the case of chronic exposure tolower doses. These effects include so-called radiation sickness causedby chronic exposure to the radiation emitting environment, and acuteradiation syndrome (poisoning), caused by acute exposure to the internalor external action of a radioactive material or a source of radiation.

Chronic exposure to low doses of radiation, particularly ionizingradiation such as gamma rays, has mutagenic activity and brings a riskof developing cancer. Harmful effects of the radiation can be also dueto the exposure of a patient or medical staff to the radiation duringroutine radiodiagnostic procedures or radiotherapy, e.g., radiotherapyof a cancer, where radiation which destroys cancer cells can at the sametime damage healthy, normal cells. This is one of the main off-targeteffects of radiotherapy of a cancer, which is urgently needed to befixed by developing effective mitigators.

Injuries to the bone marrow and gastrointestinal (GI) tract are the maindeterminants of lethality after total-body irradiation (TBI). Naturaland therapeutic protection against radiation-induced injury is mediatedby various mechanisms such as free-radical scavenging, calcium channelblockade, inhibition of lipid peroxidation, enhancement of DNA damageresponse (DDR) for effective DNA damage repair, and stimulation of stemcell and or stem-like cell activity.

Radioprotectors are compounds like anti-oxidants that can reduce thedamage in normal tissues caused by radiation only if applied before orat the time of radiation. Among them are aminothiol compounds, likemercaptamine, glutathione, amifostine, and their phosphorylatedpro-drugs. Aminothiol protectants exert their effect through theirfree-radical scavenging and antioxidant ability, and they must be givenprior to radiation exposure to provide effective protection. Adisadvantage and the main limitation of aminothiol-based radioprotectorsis their high toxicity, especially at concentrations required forradioprotection, low efficiency when used after the radiation exposure,lack of protection against radiation-induced lethality/mortality, andgenerally low degree of protection.

In contrast, mitigators may be used to minimize toxicity even afterradiation has been delivered. In recent years there has been growinginterest in biological treatments which could be administered afterradiation exposure. This includes the use of agents, e.g.,anti-apoptotic proteins, cell growth factors, G-CSF, and GM-CSF(filgrastim), which could increase survival after accidental radiationinjuries. These drugs stimulate the growth of white blood cells and canhelp repair bone marrow damage. They also can be used in patientsreceiving radiation therapy.

Due to extensive use and presence of ionizing radiation and/or radiationsources in many fields of human activity, such as medicine, nuclearpower plants, industry, as well as the threat of contamination caused bynuclear/terrorist attacks, the need still exists for radioprotectors andmitigators based on simple chemical molecules, which could be effectiveespecially in reducing radiation-induced morbidity and mortality whilebeing non-toxic and safe at concentrations required for effectiveprotection.

SUMMARY

Provided are methods useful for preventing and mitigating radiationinjury, including acute radiation syndrome. Certain embodiments of thepresent disclosure are used in preventing and mitigating radiationsyndrome, including gastrointestinal (GI) acute radiation syndrome, andpromoting overall survival following radiation injury in varioussettings, including without limitation nuclear disaster, accidentalradiation exposure, acute radiation sickness, aviation, spaceflight andspace travel, and protection and treatment of military personnel.

The present disclosure includes, for example, a method for treating orpreventing radiation-induced tissue injury, comprising administering toa subject in need thereof a therapeutically effective amount of acompound represented by formula I

or a pharmaceutically acceptable salt thereof, wherein Z is —O— or—N(H)—, thereby treating or preventing radiation-induced tissue injuryin the subject.

In certain embodiments, Z is —O— and the compound represented by formulaI is

In certain embodiments, Z is —N(H)—.

The present disclosure further includes, for example, a method fortreating or preventing radiation-induced tissue injury, comprisingadministering to a subject in need thereof a therapeutically effectiveamount of a compound represented by formula II

or a pharmaceutically acceptable salt thereof, thereby treating orpreventing radiation-induced tissue injury in the subject.

The present disclosure further includes, for example, a method fortreating or preventing radiation-induced tissue injury, comprisingadministering to a subject in need thereof a therapeutically effectiveamount of a compound represented by formula III

or a pharmaceutically acceptable salt thereof, thereby treating orpreventing radiation-induced tissue injury in the subject.

The present disclosure further includes, for example, a method fortreating or preventing radiation-induced tissue injury, comprisingco-administering to a subject in need thereof a therapeuticallyeffective amount of a compound represented by formula II

or a pharmaceutically acceptable salt thereof, and a therapeuticallyeffective amount of a compound represented by formula III

or a pharmaceutically acceptable salt thereof, thereby treating orpreventing radiation-induced tissue injury in the subject.

Accordingly, in certain embodiments of the methods disclosed herein, thesubject is a mammal.

Accordingly, in certain embodiments of the methods disclosed herein, thesubject is a human.

Accordingly, in certain embodiments of the methods disclosed herein, theradiation-induced tissue injury is prevented.

Accordingly, in certain embodiments of the methods disclosed herein, theradiation-induced tissue injury is treated.

It will be understood that the preceding summary is not to be construedas limiting the inventive concepts described in the present disclosure.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a series of photomicrographs and a related bar graph depictingthe effect of telmisartan treatment on non-tumorigenic Young AdultMurine Colonic (YAMC) epithelial cell survival in vitro after 4 Gyionizing radiation (IR). Cells in the treatment group receivedtelmisartan 24 h post IR.

FIG. 2 is a series of photomicrographs and a related bar graph depictingthe effect of telmisartan treatment on YAMC cell self-renewal in vitroafter 4 Gy IR. Cells in the treatment group received telmisartan 24 hpost IR.

FIG. 3 is a bar graph depicting in vivo whole intestinal epithelialpermeability to FITC-dextran 5 days after 14 Gy total body irradiation.Animals received three daily doses of either vehicle control ortelmisartan beginning 24 h post-IR.

FIG. 4 is a graph depicting survival of mice following 14 Gy total bodyirradiation. Animals received three daily doses of either vehiclecontrol or telmisartan beginning 24 h post-IR.

FIG. 5 is a series of photomicrographs and a related bar graph depictingthe effect of tempol treatment on YAMC cell survival in vitro after 4 GyIR. Cells in the treatment group received tempol 24 h post IR.

FIG. 6 is a series of photomicrographs and a related bar graph depictingthe effect of tempol treatment on YAMC cell self-renewal in vitro after4 Gy IR. Cells in the treatment group received tempol 24 h post IR.

FIG. 7 is a bar graph depicting in vivo whole intestinal epithelialpermeability to FITC-dextran 5 days after 14 Gy total body irradiation.Animals received three daily doses of either vehicle control or tempolbeginning 24 h post-IR.

FIG. 8 is a graph depicting survival of mice following 14 Gy total bodyirradiation. Animals received three daily doses of either vehiclecontrol or tempol beginning 24 h post-IR.

FIG. 9 is a series of photomicrographs and a related bar graph depictingthe effect of YK-4-250 treatment on YAMC cell survival in vitro after 4Gy IR. Cells in the treatment group received YK-4-250 24 h post IR.

FIG. 10 is a series of photomicrographs and a related bar graphdepicting the effect of tempol treatment on YAMC cell self-renewal invitro after 4 Gy IR. Cells in the treatment group received YK-2-250 24 hpost IR.

FIG. 11 is a bar graph depicting in vivo whole intestinal epithelialpermeability to FITC-dextran 5 days after 14 Gy total body irradiation.Animals received three daily doses of either vehicle control or YK-4-250beginning 24 h post-IR.

FIG. 12 is a graph depicting survival of mice following 14 Gy total bodyirradiation. Animals received three daily doses of either vehiclecontrol or YK-4-250 beginning 24 h post-IR.

DETAILED DESCRIPTION

Before describing various embodiments of the compounds, compositions andmethods of the present disclosure in more detail by way of exemplarydescription, examples, and results, it is to be understood that thecompounds, compositions, and methods of present disclosure are notlimited in application to the details of specific embodiments andexamples as set forth in the following description. The descriptionprovided herein is intended for purposes of illustration only and is notintended to be construed in a limiting sense. As such, the language usedherein is intended to be given the broadest possible scope and meaning;and the embodiments and examples are meant to be exemplary, notexhaustive. Also, it is to be understood that the phraseology andterminology employed herein is for the purpose of description and shouldnot be regarded as limiting unless otherwise indicated as so. Moreover,in the following detailed description, numerous specific details are setforth in order to provide a more thorough understanding of the presentdisclosure. However, it will be apparent to a person having ordinaryskill in the art that the present disclosure may be practiced withoutthese specific details. In other instances, features which are wellknown to persons of ordinary skill in the art have not been described indetail to avoid unnecessary complication of the description. It isintended that all alternatives, substitutions, modifications andequivalents apparent to those having ordinary skill in the art areincluded within the scope of the present disclosure. All of thecompounds, compositions, and methods of production and application anduse thereof disclosed herein can be made and executed without undueexperimentation in light of the present disclosure. Thus, while thecompounds, compositions, and methods of the present disclosure have beendescribed in terms of particular embodiments, it will be apparent tothose of skill in the art that variations may be applied to thecompounds, compositions and/or methods and in the steps or in thesequence of steps of the methods described herein without departing fromthe spirit, and scope of the inventive concepts.

All patents, published patent applications, and non-patent publicationsmentioned in the specification or referenced in any portion of thisapplication are herein expressly incorporated by reference in theirentirety to the same extent as if each individual patent or publicationwas specifically and individually indicated to be incorporated byreference.

Unless otherwise defined herein, scientific and technical terms used inconnection with the present disclosure shall have the meanings that arecommonly understood by those having ordinary skill in the art. Further,unless otherwise required by context, singular terms shall includepluralities and plural terms shall include the singular.

As utilized in accordance with the methods and compositions of thepresent disclosure, the following terms, unless otherwise indicated,shall be understood to have the following meanings:

The use of the word “a” or “an” when used in conjunction with the term“comprising” in the claims and/or the specification may mean “one,” butit is also consistent with the meaning of “one or more,” “at least one,”and “one or more than one.” The use of the term “or” in the claims isused to mean “and/or” unless explicitly indicated to refer toalternatives only or when the alternatives are mutually exclusive,although the disclosure supports a definition that refers to onlyalternatives and “and/or.” The use of the term “at least one” will beunderstood to include one as well as any quantity more than one,including but not limited to, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30,40, 50, 100, or any integer inclusive therein. The term “at least one”may extend up to 100 or 1000 or more, depending on the term to which itis attached; in addition, the quantities of 100/1000 are not to beconsidered limiting, as higher limits may also produce satisfactoryresults. In addition, the use of the term “at least one of X, Y and Z”will be understood to include X alone, Y alone, and Z alone, as well asany combination of X, Y and Z.

As used herein, all numerical values or ranges include fractions of thevalues and integers within such ranges and fractions of the integerswithin such ranges unless the context clearly indicates otherwise. Thus,to illustrate, reference to a numerical range, such as 1-10 includes 1,2, 3, 4, 5, 6, 7, 8, 9, 10, as well as 1.1, 1.2, 1.3, 1.4, 1.5, etc.,and so forth. Reference to a range of 1-50 therefore includes 1, 2, 3,4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, etc., upto and including 50, as well as 1.1, 1.2, 1.3, 1.4, 1.5, etc., 2.1, 2.2,2.3, 2.4, 2.5, etc., and so forth. Reference to a series of rangesincludes ranges which combine the values of the boundaries of differentranges within the series. Thus, to illustrate reference to a series ofranges, for example, of 1-10, 10-20, 20-30, 30-40, 40-50, 50-60, 60-75,75-100, 100-150, 150-200, 200-250, 250-300, 300-400, 400-500, 500-750,750-1,000, includes ranges of 1-20, 10-50, 50-100, 100-500, and500-1,000, for example.

As used in this specification and claims, the words “comprising” (andany form of comprising, such as “comprise” and “comprises”), “having”(and any form of having, such as “have” and “has”), “including” (and anyform of including, such as “includes” and “include”) or “containing”(and any form of containing, such as “contains” and “contain”) areinclusive or open-ended and do not exclude additional, unrecitedelements or method steps.

The term “or combinations thereof” as used herein refers to allpermutations and combinations of the listed items preceding the term.For example, “A, B, C, or combinations thereof” is intended to includeat least one of: A, B, C, AB, AC, BC, or ABC, and if order is importantin a particular context, also BA, CA, CB, CBA, BCA, ACB, BAC, or CAB.Continuing with this example, expressly included are combinations thatcontain repeats of one or more item or term, such as BB, AAA, AAB, BBC,AAABCCCC, CBBAAA, CABABB, and so forth. The skilled artisan willunderstand that typically there is no limit on the number of items orterms in any combination, unless otherwise apparent from the context.

Throughout this application, the term “about” is used to indicate that avalue includes the inherent variation of error for the composition, themethod used to administer the composition, or the variation that existsamong the study subjects. As used herein the qualifiers “about” or“approximately” are intended to include not only the exact value,amount, degree, orientation, or other qualified characteristic or value,but are intended to include some slight variations due to measuringerror, manufacturing tolerances, stress exerted on various parts orcomponents, observer error, wear and tear, and combinations thereof, forexample. The term “about” or “approximately”, where used herein whenreferring to a measurable value such as an amount, a temporal duration,and the like, is meant to encompass, for example, variations of ±10%, or±5%, or ±1%, or ±0.1% from the specified value, as such variations areappropriate to perform the disclosed methods and as understood bypersons having ordinary skill in the art. As used herein, the term“substantially” means that the subsequently described event orcircumstance completely occurs or that the subsequently described eventor circumstance occurs to a great extent or degree. For example, theterm “substantially” means that the subsequently described event orcircumstance occurs at least 90% of the time, or at least 95% of thetime, or at least 98% of the time.

As used herein any reference to “one embodiment” or “an embodiment”means that a particular element, feature, structure, or characteristicdescribed in connection with the embodiment is included in at least oneembodiment and may be included in other embodiments. The appearances ofthe phrase “in one embodiment” in various places in the specificationare not necessarily all referring to the same embodiment and are notnecessarily limited to a single or particular embodiment.

The term “pharmaceutically acceptable” refers to compounds andcompositions which are suitable for administration to humans and/oranimals without undue adverse side effects such as toxicity, irritationand/or allergic response commensurate with a reasonable benefit/riskratio. The compounds of the present disclosure may be combined with oneor more pharmaceutically-acceptable excipients, including carriers,vehicles, and diluents which may improve solubility, deliverability,dispersion, stability, and/or conformational integrity of the compoundsor conjugates thereof.

As used herein, “pure,” or “substantially pure” means an object speciesis the predominant species present (i.e., on a molar basis it is moreabundant than any other object species in the composition thereof), andparticularly a substantially purified fraction is a composition whereinthe object species comprises at least about 50 percent (on a molarbasis) of all macromolecular species present. Generally, a substantiallypure composition will comprise more than about 80% of all macromolecularspecies present in the composition, more particularly more than about85%, more than about 90%, more than about 95%, or more than about 99%.The term “pure” or “substantially pure” also refers to preparationswhere the object species is at least 60% (w/w) pure, or at least 70%(w/w) pure, or at least 75% (w/w) pure, or at least 80% (w/w) pure, orat least 85% (w/w) pure, or at least 90% (w/w) pure, or at least 92%(w/w) pure, or at least 95% (w/w) pure, or at least 96% (w/w) pure, orat least 97% (w/w) pure, or at least 98% (w/w) pure, or at least 99%(w/w) pure, or 100% (w/w) pure.

The term “effective amount” refers to an amount of an active agent whichis sufficient to exhibit a detectable therapeutic or treatment effect ina subject without excessive adverse side effects (such as substantialtoxicity, irritation and allergic response) commensurate with areasonable benefit/risk ratio when used in the manner of the presentdisclosure. The effective amount for a subject will depend upon thesubject's type, size and health, the nature and severity of thecondition to be treated, the method of administration, the duration oftreatment, the nature of concurrent therapy (if any), the specificformulations employed, and the like. Thus, it is not possible to specifyan exact effective amount in advance. However, the effective amount fora given situation can be determined by one of ordinary skill in the artusing routine experimentation based on the information provided herein.

The term “ameliorate” means a detectable or measurable improvement in asubject's condition or symptom thereof. A detectable or measurableimprovement includes a subjective or objective decrease, reduction,inhibition, suppression, limit or control in the occurrence, frequency,severity, progression, or duration of the condition, or an improvementin a symptom or an underlying cause or a consequence of the condition,or a reversal of the condition. A successful treatment outcome can leadto a “therapeutic effect,” or “benefit” of ameliorating, decreasing,reducing, inhibiting, suppressing, limiting, controlling or preventingthe occurrence, frequency, severity, progression, or duration of acondition, or consequences of the condition in a subject.

A decrease or reduction in worsening, such as stabilizing the condition,is also a successful treatment outcome. A therapeutic benefit thereforeneed not be complete ablation or reversal of the condition, or any one,most or all adverse symptoms, complications, consequences or underlyingcauses associated with the condition. Thus, a satisfactory endpoint maybe achieved when there is an incremental improvement such as a partialdecrease, reduction, inhibition, suppression, limit, control orprevention in the occurrence, frequency, severity, progression, orduration, or inhibition or reversal of the condition (e.g.,stabilizing), over a short or long duration of time (e.g., seconds,minutes, hours).

Telmisartan (Micardis®, Boehringer Ingelheim) is an angiotensin II (AngII) receptor antagonist (angiotensin receptor blocker; ARB) used for themanagement of hypertension. Telmisartan has high affinity for theangiotensin II receptor type 1 (AT₁), with a binding affinity 3000 timesgreater for AT₁ than for AT₂. It has the longest half-life of any ARB(24 hours) and the largest volume of distribution among ARBs (500liters). In addition to blocking the angiotensin receptors, telmisartanacts as a selective modulator of peroxisome proliferator-activatedreceptor gamma (PPAR-γ), a central regulator of insulin and glucosemetabolism. It is believed that telmisartan's dual mode of action mayprovide protective benefits against the vascular and renal damage causedby diabetes and cardiovascular disease.

Telmisartan is represented by formula II

Tempol or 4-Hydroxy-TEMPO, formally4-hydroxy-2,2,6,6-tetramethylpiperidin-1-oxyl, is a heterocycliccompound. Like the related TEMPO((2,2,6,6-tetramethylpiperidin-1-yl)oxyl), it is used as a catalyst andchemical oxidant by virtue of being a stable radical. Tempol isrecognized as an agent for detoxifying reactive oxygen species. Itcatalyzes the disproportionation of superoxide, facilitates hydrogenperoxide metabolism, and inhibit Fenton chemistry. Tempol iscommercially available from any of a number of suppliers, e.g.,Sigma-Aldrich.

Tempol is represented by formula III

U.S. Pat. No. 9,233,949 to Brown et al. discloses an ester-linkedconjugate formed between telmisartan and tempol. This conjugate,designated YK-4-250, is represented by:

YK-4-250 is disclosed in U.S. Pat. No. 9,233,949 to be useful to treatoxidative stress and/or hypertension by reducing Ang II-stimulatedvascular superoxide (O₂·⁻) and blood pressure.

Exposure of living cells and tissue to high amounts of ionizingradiation causes physical damage or injury to cells and tissues. Thedamage or injury can include damage to DNA

Acute radiation syndrome is typically caused by exposure to a large doseof ionizing radiation, e.g., for humans, greater than or equal to about0.1 Gray (Gy), over a short period of time, e.g., one hour. Morbidityand mortality increase with absorbed dose. Acute radiation syndrome isalso known as radiation poisoning, radiation sickness, and radiationtoxicity.

Symptoms of acute radiation syndrome typically can include any one ormore of nausea, vomiting, diarrhea, fever, headache, cognitiveimpairment, seizures, tremor, ataxia, lethargy, infection, anemia, andbleeding. Historically, at 6-8 Gy, mortality is 50-100 percent even withtreatment, while at greater than 8 Gy, mortality is 100 percent evenwith treatment.

Methods

In at least one embodiment, the present disclosure is directed to amethod for treating or preventing radiation-induced tissue injury,comprising administering to a subject in need thereof a therapeuticallyeffective amount of a compound represented by formula I

or a pharmaceutically acceptable salt thereof, wherein Z is —O— or—N(H)—, thereby treating or preventing radiation-induced tissue injuryin the subject.

In certain embodiments, Z is —O— and the compound represented by formulaI is

In certain embodiments, Z is —N(H)—.

As used herein, “treat” means to reduce or mitigate at least one sign orsymptom of an existing disease or condition of a subject. In certainembodiments, “treat” means to reduce or mitigate the severity of atleast one sign or symptom of an existing disease or condition of asubject. In certain embodiments, “treat” means to reduce the duration ofat least one sign or symptom of an existing disease or condition of asubject. In certain embodiments, “treat” means to resolve at least onesign or symptom of an existing disease or condition of a subject. Incertain embodiments, “treat” means to restore the health of a subject,e.g., to restore to the health of the subject prior to the subject'sdevelopment of the disease or condition. As used herein, “prevent” or“prevention” refers to prophylactic treatment measures to stop the onsetof a condition or disease.

As used herein, “subject” refers to a living mammal. In certainembodiments, the subject is a mouse, rat, hamster, guinea pig, rabbit,sheep, goat, cat, dog, horse, cow, or non-human primate. In certainembodiments, the subject is a human.

As used herein, “radiation-induced tissue injury” refers to any injuryor damage to cells of a tissue, where such injury or damage isconsistent with or attributable to known exposure of the cells toionizing radiation. The term “radiation-induced tissue injury” can referto injury involving any one or more tissues, including acute radiationsyndrome.

In certain embodiments, the radiation-induced tissue injury isprevented.

In certain embodiments, the radiation-induced tissue injury is treated.

In certain embodiments, the compound is administered prior to exposureof the subject to radiation. Such administration of the compound maythen prevent or treat radiation-induced tissue injury in the subject.The agent administered in accordance with such embodiments can act bothas a radioprotector and as a mitigator. For example, a subject about toembark on spaceflight or about to encounter a situation or environmentknown or believed to carry the risk of exposure to ionizing radiationmay be administered the compound prior to said spaceflight or prior toencountering said situation or environment. The method is useful insettings including, but not limited to, nuclear disaster, accidentalradiation exposure, acute radiation sickness, aviation, space travel,and protection of military personnel.

In certain embodiments, the compound is administered during exposure ofthe subject to radiation. Such administration of the compound may thenprevent or treat radiation-induced tissue injury in the subject. Theagent administered in accordance with such embodiments acts as aradioprotector and/or as a mitigator. The method is useful in settingsincluding, but not limited to, nuclear disaster, accidental radiationexposure, acute radiation sickness, aviation, spaceflight and spacetravel, and treatment of military personnel.

In certain embodiments, the compound is administered after exposure ofthe subject to radiation. Such administration of the compound thentreats radiation-induced tissue injury in the subject. The agentadministered in accordance with such embodiments acts as a mitigator.The method is useful in settings including, but not limited to, nucleardisaster, accidental radiation exposure, acute radiation sickness,aviation, spaceflight and space travel, and treatment of militarypersonnel.

In certain embodiments, the radiation-induced tissue injury is acuteradiation-induced tissue injury.

In certain embodiments, the radiation-induced tissue injury is chronicradiation-induced tissue injury.

In certain embodiments, the tissue comprises gastrointestinal tissue. Incertain embodiments, the tissue is gastrointestinal tissue. As usedherein, “gastrointestinal tissue” refers generally to mouth, pharynx,esophagus, stomach, small intestine, large intestine, and rectum,including in particular epithelia lining said structures. In certainembodiments, “gastrointestinal tissue” refers to mouth, including inparticular epithelia lining the mouth. In certain embodiments,“gastrointestinal tissue” refers to pharynx, including in particularepithelia lining the pharynx. In certain embodiments, “gastrointestinaltissue” refers to esophagus, including in particular epithelia liningthe esophagus. In certain embodiments, “gastrointestinal tissue” refersto stomach, including in particular epithelia lining the stomach. Incertain embodiments, “gastrointestinal tissue” refers to smallintestine, including in particular epithelia lining the small intestine.In certain embodiments, “gastrointestinal tissue” refers to largeintestine, including in particular epithelia lining the large intestine.In certain embodiments, “gastrointestinal tissue” refers to rectum,including in particular epithelia lining the rectum.

In certain embodiments, the tissue comprises hematopoietic tissue. Incertain embodiments, the tissue is hematopoietic tissue. As used herein,“hematopoietic tissue” refers generally to blood cell-forming componentsof bone marrow. In certain embodiments, “hematopoietic tissue” refers tored blood cell-forming components of bone marrow. In certainembodiments, “hematopoietic tissue” refers to white blood cell-formingcomponents of bone marrow. In certain embodiments, “hematopoietictissue” also refers to thymus. In certain embodiments, “hematopoietictissue” also refers to lymph nodes. In certain embodiments,“hematopoietic tissue” also refers to spleen. In certain embodiments,“hematopoietic tissue” also refers to liver.

In certain embodiments, the tissue comprises neural tissue. In certainembodiments, the tissue is neural tissue. As used herein, “neuraltissue” refers generally to neurons of the central nervous system andperipheral nerves. In certain embodiments, “neural tissue” refers toneurons of the central nervous system. In certain embodiments, “neuraltissue” refers to neurons of the brain. In certain embodiments, “neuraltissue” refers to neurons of the spinal cord. In certain embodiments,“neural tissue” refers to neurons of peripheral nerves.

In certain embodiments, the tissue is selected from the group consistingof skin, eye, thyroid, heart, lung, kidney, adrenal gland, pancreas,gall bladder, breast, prostate, ovary, testis, uterus, vagina, andvascular tissue.

In certain embodiments, the compound is administered orally.

In certain embodiments, the compound is administered parenterally.

In at least one embodiment, the present disclosure is directed to amethod for treating or preventing radiation-induced tissue injury,comprising administering to a subject in need thereof a therapeuticallyeffective amount of a compound represented by formula II (telmisartan)

or a pharmaceutically acceptable salt thereof, thereby treating orpreventing radiation-induced tissue injury in the subject.

In certain embodiments, the subject is a mouse, rat, hamster, guineapig, rabbit, sheep, goat, cat, dog, horse, cow, or non-human primate. Incertain embodiments, the subject is a human.

In certain embodiments, the radiation-induced tissue injury isprevented.

In certain embodiments, the radiation-induced tissue injury is treated.

In certain embodiments, the compound is administered prior to exposureof the subject to radiation. Such administration of the compound maythen prevent or treat radiation-induced tissue injury in the subject.The administered agent in accordance with such embodiments can act bothas a radioprotector and as a mitigator. For example, a subject about toembark on spaceflight or about to encounter a situation or environmentknown or believed to carry the risk of exposure to ionizing radiationmay be administered the compound prior to said spaceflight orencountering said situation or environment. The method is useful insettings including, but not limited to, nuclear disaster, accidentalradiation exposure, acute radiation sickness, aviation, space travel,and protection of military personnel.

In certain embodiments, the compound is administered during exposure ofthe subject to radiation. Such administration of the compound may thenprevent or treat radiation-induced tissue injury in the subject. Theagent administered in accordance with such embodiments acts as aradioprotector and/or as a mitigator. The method is useful in settingsincluding, but not limited to, nuclear disaster, accidental radiationexposure, acute radiation sickness, aviation, spaceflight and spacetravel, and treatment of military personnel.

In certain embodiments, the compound is administered after exposure ofthe subject to radiation. Such administration of the compound thentreats radiation-induced tissue injury in the subject. The administeredagent in accordance with such embodiments acts as a mitigator. Themethod is useful in settings including, but not limited to, nucleardisaster, accidental radiation exposure, acute radiation sickness,aviation, spaceflight and space travel, and treatment of militarypersonnel.

In certain embodiments, the radiation-induced tissue injury is acuteradiation-induced tissue injury.

In certain embodiments, the radiation-induced tissue injury is chronicradiation-induced tissue injury.

In certain embodiments, the tissue comprises gastrointestinal tissue. Incertain embodiments, the tissue is gastrointestinal tissue. In certainembodiments, “gastrointestinal tissue” refers to mouth, including inparticular epithelia lining the mouth. In certain embodiments,“gastrointestinal tissue” refers to pharynx, including in particularepithelia lining the pharynx. In certain embodiments, “gastrointestinaltissue” refers to esophagus, including in particular epithelia liningthe esophagus. In certain embodiments, “gastrointestinal tissue” refersto stomach, including in particular epithelia lining the stomach. Incertain embodiments, “gastrointestinal tissue” refers to smallintestine, including in particular epithelia lining the small intestine.In certain embodiments, “gastrointestinal tissue” refers to largeintestine, including in particular epithelia lining the large intestine.In certain embodiments, “gastrointestinal tissue” refers to rectum,including in particular epithelia lining the rectum.

In certain embodiments, the tissue comprises hematopoietic tissue. Incertain embodiments, the tissue is hematopoietic tissue. In certainembodiments, “hematopoietic tissue” refers to red blood cell-formingcomponents of bone marrow. In certain embodiments, “hematopoietictissue” refers to white blood cell-forming components of bone marrow. Incertain embodiments, “hematopoietic tissue” also refers to thymus. Incertain embodiments, “hematopoietic tissue” also refers to lymph nodes.In certain embodiments, “hematopoietic tissue” also refers to spleen. Incertain embodiments, “hematopoietic tissue” also refers to liver.

In certain embodiments, the tissue comprises neural tissue. In certainembodiments, the tissue is neural tissue. In certain embodiments,“neural tissue” refers to neurons of the central nervous system. Incertain embodiments, “neural tissue” refers to neurons of the brain. Incertain embodiments, “neural tissue” refers to neurons of the spinalcord. In certain embodiments, “neural tissue” refers to neurons ofperipheral nerves.

In certain embodiments, the tissue is selected from the group consistingof skin, eye, thyroid, heart, lung, kidney, adrenal gland, pancreas,gall bladder, breast, prostate, ovary, testis, uterus, vagina, andvascular tissue.

In certain embodiments, the compound is administered orally.

In certain embodiments, the compound is administered parenterally.

In at least one embodiment, the present disclosure is directed to amethod for treating or preventing radiation-induced tissue injury,comprising administering to a subject in need thereof a therapeuticallyeffective amount of a compound represented by formula III (tempol)

or a pharmaceutically acceptable salt thereof, to treat or preventradiation-induced tissue injury in the subject.

In certain embodiments, the subject is a mouse, rat, hamster, guineapig, rabbit, sheep, goat, cat, dog, horse, cow, or non-human primate. Incertain embodiments, the subject is a human.

In certain embodiments, the radiation-induced tissue injury isprevented.

In certain embodiments, the radiation-induced tissue injury is treated.

In certain embodiments, the compound is administered prior to exposureof the subject to radiation. Such administration of the compound maythen prevent or treat radiation-induced tissue injury in the subject.The administered agent in accordance with such embodiments can act bothas a radioprotector and as a mitigator. For example, a subject about toembark on spaceflight or about to encounter a situation or environmentknown or believed to carry the risk of exposure to ionizing radiationmay be administered the compound prior to said spaceflight orencountering said situation or environment. The method is useful insettings including, but not limited to, nuclear disaster, accidentalradiation exposure, acute radiation sickness, aviation, space travel,and protection of military personnel.

In certain embodiments, the compound is administered during exposure ofthe subject to radiation. Such administration of the compound may thenprevent or treat radiation-induced tissue injury in the subject. Theagent administered in accordance with such embodiments acts as aradioprotector and/or as a mitigator. The method is useful in settingsincluding, but not limited to, nuclear disaster, accidental radiationexposure, acute radiation sickness, aviation, spaceflight and spacetravel, and treatment of military personnel.

In certain embodiments, the compound is administered after exposure ofthe subject to radiation. Such administration of the compound thentreats radiation-induced tissue injury in the subject. The administeredagent in accordance with such embodiments acts as a mitigator. Themethod is useful in settings including, but not limited to, nucleardisaster, accidental radiation exposure, acute radiation sickness,aviation, spaceflight and space travel, and treatment of militarypersonnel.

In certain embodiments, the radiation-induced tissue injury is acuteradiation-induced tissue injury.

In certain embodiments, the radiation-induced tissue injury is chronicradiation-induced tissue injury.

In certain embodiments, the tissue comprises gastrointestinal tissue. Incertain embodiments, the tissue is gastrointestinal tissue. In certainembodiments, “gastrointestinal tissue” refers to mouth, including inparticular epithelia lining the mouth. In certain embodiments,“gastrointestinal tissue” refers to pharynx, including in particularepithelia lining the pharynx. In certain embodiments, “gastrointestinaltissue” refers to esophagus, including in particular epithelia liningthe esophagus. In certain embodiments, “gastrointestinal tissue” refersto stomach, including in particular epithelia lining the stomach. Incertain embodiments, “gastrointestinal tissue” refers to smallintestine, including in particular epithelia lining the small intestine.In certain embodiments, “gastrointestinal tissue” refers to largeintestine, including in particular epithelia lining the large intestine.In certain embodiments, “gastrointestinal tissue” refers to rectum,including in particular epithelia lining the rectum.

In certain embodiments, the tissue comprises hematopoietic tissue. Incertain embodiments, the tissue is hematopoietic tissue. In certainembodiments, “hematopoietic tissue” refers to red blood cell-formingcomponents of bone marrow. In certain embodiments, “hematopoietictissue” refers to white blood cell-forming components of bone marrow. Incertain embodiments, “hematopoietic tissue” also refers to thymus. Incertain embodiments, “hematopoietic tissue” also refers to lymph nodes.In certain embodiments, “hematopoietic tissue” also refers to spleen. Incertain embodiments, “hematopoietic tissue” also refers to liver.

In certain embodiments, the tissue comprises neural tissue. In certainembodiments, the tissue is neural tissue. In certain embodiments,“neural tissue” refers to neurons of the central nervous system. Incertain embodiments, “neural tissue” refers to neurons of the brain. Incertain embodiments, “neural tissue” refers to neurons of the spinalcord. In certain embodiments, “neural tissue” refers to neurons ofperipheral nerves.

In certain embodiments, the tissue is selected from the group consistingof skin, eye, thyroid, heart, lung, kidney, adrenal gland, pancreas,gall bladder, breast, prostate, ovary, testis, uterus, vagina, andvascular tissue.

In certain embodiments, the compound is administered orally.

In certain embodiments, the compound is administered parenterally.

In at least one embodiment, the present disclosure is directed to amethod for treating or preventing radiation-induced tissue injury,comprising co-administering to a subject in need thereof atherapeutically effective amount of a compound represented by formula II

or a pharmaceutically acceptable salt thereof, and a therapeuticallyeffective amount of a compound represented by formula III

or a pharmaceutically acceptable salt thereof, thereby treating orpreventing radiation-induced tissue injury in the subject.

As used herein, “co-administering” refers to administering a pluralityof agents to a subject such that each of the plurality of agents ispresent in the subject simultaneously. In certain embodiments, agentsare included in a single dosage form. In certain embodiments, the agentsare administered simultaneously in separate dosage forms. In certainembodiments, the agents are administered sequentially in separate dosageforms.

In certain embodiments, the subject is a mouse, rat, hamster, guineapig, rabbit, sheep, goat, cat, dog, horse, cow, or non-human primate. Incertain embodiments, the subject is a human.

In certain embodiments, the radiation-induced tissue injury isprevented.

In certain embodiments, the radiation-induced tissue injury is treated.

In certain embodiments, the compounds are co-administered prior toexposure of the subject to radiation. Such co-administration of thecompounds may then prevent or treat radiation-induced tissue injury inthe subject. The co-administered agents in accordance with suchembodiments can act both as radioprotectors and as mitigators. Forexample, a subject about to embark on spaceflight or about to encountera situation or environment known or believed to carry the risk ofexposure to ionizing radiation may be co-administered the compoundsprior to said spaceflight or encountering said situation or environment.The method is useful in settings including, but not limited to, nucleardisaster, accidental radiation exposure, acute radiation sickness,aviation, space travel, and protection of military personnel.

In certain embodiments, the compounds are co-administered duringexposure of the subject to radiation. Such co-administration of thecompounds may then prevent or treat radiation-induced tissue injury inthe subject. The agents co-administered in accordance with suchembodiments act as radioprotectors and/or as mitigators. The method isuseful in settings including, but not limited to, nuclear disaster,accidental radiation exposure, acute radiation sickness, aviation,spaceflight and space travel, and treatment of military personnel.

In certain embodiments, the compounds are co-administered after exposureof the subject to radiation. Such co-administration of the compoundsthen treats radiation-induced tissue injury in the subject. Theco-administered agents in accordance with such embodiments act asmitigators. The method is useful in settings including, but not limitedto, nuclear disaster, accidental radiation exposure, acute radiationsickness, aviation, spaceflight and space travel, and treatment ofmilitary personnel.

In certain embodiments, the radiation-induced tissue injury is acuteradiation-induced tissue injury.

In certain embodiments, the radiation-induced tissue injury is chronicradiation-induced tissue injury.

In certain embodiments, the tissue comprises gastrointestinal tissue. Incertain embodiments, the tissue is gastrointestinal tissue. In certainembodiments, “gastrointestinal tissue” refers to mouth, including inparticular epithelia lining the mouth. In certain embodiments,“gastrointestinal tissue” refers to pharynx, including in particularepithelia lining the pharynx. In certain embodiments, “gastrointestinaltissue” refers to esophagus, including in particular epithelia liningthe esophagus. In certain embodiments, “gastrointestinal tissue” refersto stomach, including in particular epithelia lining the stomach. Incertain embodiments, “gastrointestinal tissue” refers to smallintestine, including in particular epithelia lining the small intestine.In certain embodiments, “gastrointestinal tissue” refers to largeintestine, including in particular epithelia lining the large intestine.In certain embodiments, “gastrointestinal tissue” refers to rectum,including in particular epithelia lining the rectum.

In certain embodiments, the tissue comprises hematopoietic tissue. Incertain embodiments, the tissue is hematopoietic tissue. In certainembodiments, “hematopoietic tissue” refers to red blood cell-formingcomponents of bone marrow. In certain embodiments, “hematopoietictissue” refers to white blood cell-forming components of bone marrow. Incertain embodiments, “hematopoietic tissue” also refers to thymus. Incertain embodiments, “hematopoietic tissue” also refers to lymph nodes.In certain embodiments, “hematopoietic tissue” also refers to spleen. Incertain embodiments, “hematopoietic tissue” also refers to liver.

In certain embodiments, the tissue comprises neural tissue. In certainembodiments, the tissue is neural tissue. In certain embodiments,“neural tissue” refers to neurons of the central nervous system. Incertain embodiments, “neural tissue” refers to neurons of the brain. Incertain embodiments, “neural tissue” refers to neurons of the spinalcord. In certain embodiments, “neural tissue” refers to neurons ofperipheral nerves.

In certain embodiments, the tissue is selected from the group consistingof skin, eye, thyroid, heart, lung, kidney, adrenal gland, pancreas,gall bladder, breast, prostate, ovary, testis, uterus, vagina, andvascular tissue.

In certain embodiments, the compounds are co-administered orally.

In certain embodiments, the compounds are co-administered parenterally.

In certain embodiments, the compounds are co-administered sequentially.

In certain embodiments, the compounds are co-administered substantiallysimultaneously.

Formulation and Dosing

Suitable routes of administration include, but not limited to, oral andparenteral. Suitable routes of parenteral administration may, forexample, include intravenous, intraperitoneal, intramuscular,subcutaneous, rectal, transmucosal, topical, pulmonary, and intrathecal.

Alternatively, administration may be in a local rather than a systemicmanner, for example, via injection of a compound directly into aspecific anatomical site, often in a depot or sustained releaseformulation.

Furthermore, the administration may be in a targeted drug deliverysystem, for example, in a liposome coated with cell-specific antibody orother targeting agent.

The pharmaceutical compositions of the present disclosure may bemanufactured in a manner that is itself known, e.g., by means ofconventional mixing, dissolving, granulating, dragee-making, levigating,emulsifying, encapsulating, entrapping or lyophilizing processes.

Pharmaceutical compositions for use in accordance with the presentdisclosure thus may be formulated in a conventional manner using one ormore physiologically acceptable carriers comprising excipients andauxiliaries which facilitate processing of the active compounds intopreparations which can be used pharmaceutically. Proper formulation isdependent upon the route of administration chosen.

For injection, the active agent or agents used in the methods of thepresent disclosure may be formulated in aqueous solutions, preferably inphysiologically compatible buffers such as Hanks' solution, Ringer'ssolution, or physiological saline buffer. For transmucosaladministration, penetrants appropriate to the barrier to be permeatedare used in the formulation. Such penetrants are generally known in theart.

For oral administration, the active agent or agents can be formulatedreadily by combining the active compounds with pharmaceuticallyacceptable carriers well known in the art. Such carriers enable themixtures or adducts of the invention to be formulated as tablets, pills,dragees, capsules, liquids, gels, syrups, slurries, suspensions and thelike, for oral ingestion by a subject to be treated. Pharmaceuticalpreparations for oral use can be obtained by combining the mixtures oradducts of the invention with a solid excipient, optionally grinding aresulting mixture, and processing the mixture of granules, after addingsuitable auxiliaries, if desired, to obtain tablets or dragee cores.Suitable excipients are, in particular, fillers such as sugars,including lactose, sucrose, mannitol, or sorbitol; cellulosepreparations such as, for example, maize starch, wheat starch, ricestarch, potato starch, gelatin, gum tragacanth, methyl cellulose,hydroxypropylmethyl-cellulose, sodium carboxymethylcellulose, and/orpolyvinylpyrrolidone (PVP). If desired, disintegrating agents may beadded, such as the cross-linked polyvinyl pyrrolidone, agar, or alginicacid or a salt thereof such as sodium alginate.

In one embodiment the pharmaceutically acceptable carrier excludesdimethylsulfoxide (DMSO).

Dragee cores are provided with suitable coatings. For this purpose,concentrated sugar solutions may be used, which may optionally containgum arabic, talc, polyvinyl pyrrolidone, carbopol gel, polyethyleneglycol, and/or titanium dioxide, lacquer solutions, and suitable organicsolvents or solvent mixtures. Dyestuffs or pigments may be added to thetablets or dragee coatings for identification or to characterizedifferent combinations of active compound doses.

Pharmaceutical preparations which can be used orally include push-fitcapsules made of gelatin, as well as soft, sealed capsules made ofgelatin and a plasticizer, such as glycerol or sorbitol. The push-fitcapsules can contain the active ingredients in admixture with fillersuch as lactose, binders such as starches, and/or lubricants such astalc or magnesium stearate and, optionally, stabilizers. In softcapsules, the active compounds may be dissolved or suspended in suitableliquids, such as fatty oils, liquid paraffin, or liquid polyethyleneglycols. In addition, stabilizers may be added. All formulations fororal administration should be in dosages suitable for suchadministration.

For buccal administration, the compositions may take the form of tabletsor lozenges formulated in conventional manner.

For administration by inhalation, the active agent or agents for useaccording to the present disclosure are conveniently delivered in theform of an aerosol spray presentation from pressurized packs or anebuliser, with the use of a suitable propellant, e.g.,dichlorodifluoromethane, trichlorofluoromethane,dichlorotetrafluoroethane, carbon dioxide or other suitable gas. In thecase of pressurized aerosol the dosage unit may be determined byproviding a valve to deliver a metered amount. Capsules and cartridgesof e.g., gelatin for use in an inhaler or insufflator may be formulatedcontaining a powder mix of the compound and a suitable powder base suchas lactose or starch.

The active agent or agents of the invention can be formulated forparenteral administration by injection, e.g., bolus injection orcontinuous infusion. Formulations for injection may be presented in unitdosage form, e.g., in ampoules or in multi-dose containers, with anadded preservative. The compositions may take such forms as suspensions,solutions or emulsions in oily or aqueous vehicles, and may containformulatory agents such as suspending, stabilizing and/or dispersingagents.

Pharmaceutical formulations for parenteral administration includeaqueous solutions of the active compounds in water-soluble form.Additionally, suspensions of the mixtures or adducts of the presentdisclosure may be prepared as appropriate oily injection suspensions.Suitable lipophilic solvents or vehicles include fatty oils such assesame oil, or synthetic fatty acid esters, such as ethyl oleate ortriglycerides, or liposomes. Aqueous injection suspensions may containsubstances which increase the viscosity of the suspension, such assodium carboxymethyl cellulose, sorbitol, or dextran. Optionally, thesuspension may also contain suitable stabilizers or agents whichincrease the solubility of the compounds to allow for the preparation ofhighly concentrated solutions.

Alternatively, the active agent or agents of the present disclosure maybe in powder form for constitution with a suitable vehicle, e.g.,sterile pyrogen-free water, before use.

The active agent or agents of the present disclosure may also beformulated in rectal compositions such as suppositories or retentionenemas, e.g., containing conventional suppository bases such as cocoabutter or other glycerides.

In addition to the formulations described previously, the active agentor agents of the present disclosure may also be formulated as a depotpreparation. Such long acting formulations may be administered byimplantation (for example subcutaneously or intramuscularly or byintramuscular injection). Thus, for example, the mixtures or adducts ofthe present disclosure may be formulated with suitable polymeric orhydrophobic materials (for example as an emulsion in an acceptable oil)or ion exchange resins, or as sparingly soluble derivatives, forexample, as a sparingly soluble salt.

Alternatively, other delivery systems for hydrophobic pharmaceuticalcompounds may be employed. Liposomes and emulsions are well knownexamples of delivery vehicles or carriers for hydrophobic drugs. Certainorganic solvents such as dimethysulfoxide (DMSO) also may be employed,although usually at the cost of greater toxicity. Additionally, thecompounds may be delivered using a sustained-release system, such assemipermeable matrices of solid hydrophobic polymers containing thetherapeutic agent. Various sustained-release materials have beenestablished and are well known by those skilled in the art.Sustained-release capsules may, depending on their chemical nature,release the compounds for a few weeks up to over 100 days. Depending onthe chemical nature and the biological stability of the therapeuticreagent, additional strategies for protein stabilization may beemployed.

The pharmaceutical compositions also may comprise suitable solid or gelphase carriers or excipients. Examples of such carriers or excipientsinclude but are not limited to calcium carbonate, calcium phosphate,various sugars, starches, cellulose derivatives, gelatin, and polymerssuch as polyethylene glycols.

Compounds for use according to the present disclosure may be provided assalts with pharmaceutically compatible counterions (i.e.,pharmaceutically acceptable salts). A “pharmaceutically acceptable salt”means any non-toxic salt that, upon administration to a recipient, iscapable of providing, either directly or indirectly, a compound or aprodrug of a compound of this invention. A “pharmaceutically acceptablecounterion” is an ionic portion of a salt that is not toxic whenreleased from the salt upon administration to a recipient.Pharmaceutically compatible salts may be formed with many acids,including but not limited to hydrochloric, sulfuric, acetic, lactic,tartaric, malic, succinic, etc. Salts tend to be more soluble in aqueousor other protonic solvents than are the corresponding free base forms.

Acids commonly employed to form pharmaceutically acceptable saltsinclude inorganic acids such as hydrogen bisulfide, hydrochloric,hydrobromic, hydroiodic, sulfuric and phosphoric acid, as well asorganic acids such as para-toluenesulfonic, salicylic, tartaric,bitartaric, ascorbic, maleic, besylic, fumaric, gluconic, glucuronic,formic, glutamic, methanesulfonic, ethanesulfonic, benzenesulfonic,lactic, oxalic, para-bromophenylsulfonic, carbonic, succinic, citric,benzoic and acetic acid, and related inorganic and organic acids. Suchpharmaceutically acceptable salts thus include sulfate, pyrosulfate,bisulfate, sulfite, bisulfite, phosphate, monohydrogenphosphate,dihydrogenphosphate, metaphosphate, pyrophosphate, chloride, bromide,iodide, acetate, propionate, decanoate, caprylate, acrylate, formate,isobutyrate, caprate, heptanoate, propiolate, oxalate, malonate,succinate, suberate, sebacate, fumarate, maleate, butyne-1,4-dioate,hexyne-1,6-dioate, benzoate, chlorobenzoate, methylbenzoate,dinitrobenzoate, hydroxybenzoate, methoxybenzoate, phthalate,terephathalate, sulfonate, xylenesulfonate, phenylacetate,phenylpropionate, phenylbutyrate, citrate, lactate,.beta.-hydroxybutyrate, glycolate, maleate, tartrate, methanesulfonate,propanesulfonate, naphthalene-1-sulfonate, naphthalene-2-sulfonate,mandelate and the like salts. Preferred pharmaceutically acceptable acidaddition salts include those formed with mineral acids such ashydrochloric acid and hydrobromic acid, and especially those formed withorganic acids such as maleic acid.

Suitable bases for forming pharmaceutically acceptable salts with acidicfunctional groups include, but are not limited to, hydroxides of alkalimetals such as sodium, potassium, and lithium; hydroxides of alkalineearth metal such as calcium and magnesium; hydroxides of other metals,such as aluminum and zinc; ammonia, and organic amines, such asunsubstituted or hydroxy-substituted mono-, di-, or trialkylamines;dicyclohexylamine; tributyl amine; pyridine; N-methyl-N-ethylamine;diethylamine; triethylamine; mono-, bis-, or tris-(2-hydroxy-lower alkylamines), such as mono-, bis-, or tris-(2-hydroxyethyl)amine,2-hydroxy-tert-butylamine, or tris-(hydroxymethyl)methylamine, N,N-dialkyl-N-(hydroxy alkyl)-amines, such asN,N-dimethyl-N-(2-hydroxyethyl)amine, or tri-(2-hydroxyethyl)amine;N-methyl-D-glucamine; and amino acids such as arginine, lysine, and thelike.

Pharmaceutical compositions suitable for use in the present disclosureinclude compositions wherein each active agent is contained in aneffective amount to achieve its intended purpose.

As used herein, a “therapeutically effective amount” is an amount whichinhibits, totally or partially, the progression of a condition to betreated; or alleviates, at least partially, one or more symptoms of saidcondition. Alternatively or in addition, in certain embodiments atherapeutically effective amount can be an amount which isprophylactically effective. The amount which is therapeuticallyeffective will depend upon factors such as the subject's size andgender, the condition to be treated, the severity of the condition andthe result sought. For a given subject, a therapeutically effectiveamount can be determined by methods known to those of skill in the art.

In the methods of the invention the active agent or agents can beadministered orally or by injection, using any pharmaceutical dosageforms known for the person skilled in the art for such administration.Dosing may be a single dose or multiple doses.

A manner of administration is oral administration.

For oral administration both solid and liquid formulations can be used.Solid formulations include conventional tablets, capsules, troches,powders or granulates for direct ingestion or for reconstitution inliquids, such as water or juices. Any suitable conventional excipientscan be used for the preparation of such solid forms. Liquid forms fororal administration include in particular aqueous solutions, with theaddition of any conventional excipients, such as for example flavorsand/or sweeteners.

Another manner of administration is parenteral administration,especially by continuous infusion or a single bolus.

Administration by injection includes bolus intravenous injection,continuous intravenous infusion, as well as subcutaneous injection. Anysuitable injection formulation can be used, such as for example aqueoussaline solutions, buffered saline solutions, etc., using conventionalexcipients known from the art, such as preservatives, isotonic agents,buffers, etc.

The administration, in a single dose or in multiple doses, can be priorto or after a radiation exposure. In the case of multiple doses,administration can be continued in cycles, such as every day, everyother day, or every several days. The administration can be continuedfor several days or weeks. In the case of long-term radiation exposure,the administration can be during the whole exposure period and can becontinued after cessation of the exposure for several days or weeks.

Dosage can be based, at least in part, on results of in vitro testingand/or in vivo testing in laboratory animals, some methods for which aredescribed in the examples below.

Actual dosage levels of the active agent or agents in the methods of thepresent disclosure may be varied so as to obtain an amount of the activeingredient which is effective to achieve the desired therapeuticresponse for a particular patient, composition, and mode ofadministration, preferably without being toxic to the patient.

Dosage regimen in the case of the administration as a protective measurein the case of routine, expected, or predictable long-term exposure tolow-dose ionizing radiation can be in the range of about 1 mg/kg bodyweight to 100 mg/kg body weight of the active agent or agents per day.The active agent or agents can be administered as a protective measurebefore the onset to the exposure in one single dose, or alternatively individed doses, during the period of such exposure and optionally forsome period after cessation of the exposure. The administration can bepreferably repeated daily.

In the case of expected acute exposure, such as for example the exposureof a professional rescue or emergency staff after accidents involvingionizing radiation, the active agent or agents should be administeredprior the expected exposure, in one single dose or in multiple doses andpreferably its administration should be continued for the whole periodof exposure.

Dosage regimens in the case of administration as a part of aradiotherapy procedure can be in, but are not limited to, the range ofabout 2 mg/kg body weight to 200 mg/kg body weight of the active agentor agents in a single dose before irradiation, such as, but not limitedto, 1 hour, 30 minutes, or 15 minutes before irradiation. In particular,dosages such as 5 mg/kg to 20 mg/kg can be used, or 5 mg/kg to 10 mg/kg.In certain embodiments, a dose of the active agent or agents is about 1mg/kg, about 2 mg/kg, about 3 mg/kg, about 4 mg/kg, about 5 mg/kg, about6 mg/kg, about 7 mg/kg, about 8 mg/kg, about 9 mg/kg, about 10 mg/kg,about 11 mg/kg, about 12 mg/kg, about 13 mg/kg, about 14 mg/kg, about 15mg/kg, about 16 mg/kg, about 17 mg/kg, about 18 mg/kg, about 19 mg/kg,or about 20 mg/kg.

In the case of accidental acute exposure, the active agent or agentsshould be administered to the victim of such accidental exposure as soonas possible after the exposure has taken place and typically should becontinued daily for some period, such as several days or weeks, afterthe exposure. In such cases a dosage regimen of about, but not limitedto, 2 mg/kg body weight to about 200 mg/kg body weight per day iscontemplated, depending on the radiation dose absorbed by the subject,the route of administration, and/or the clinical condition of thesubject.

Generally, the dose of the active agent or agents will vary depending onthe absorbed or expected dose of the radiation. The selected dosagelevel will also depend upon a variety of factors including the activityof the particular active agent or agents employed, the time ofadministration, the rate of excretion of the particular active agent oragents being employed, the duration of the treatment, other drugs,compounds or materials used in combination with the particular activeagent or agents, the age, sex, weight, condition, general health, andprior medical history of the patient being treated, and like factorswell known in the medical arts.

A medical doctor, e.g., physician or veterinarian, having ordinary skillin the art can readily determine and prescribe the effective amount ofthe active agent or agents required. A medical doctor, e.g., physicianor veterinarian, having ordinary skill in the art can readily determineand prescribe the effective amount of a pharmaceutical composition orpharmaceutical compositions comprising the active agent or agentsrequired.

One of ordinary skill in the art would be able to study the relevantfactors and make the determination regarding the effective amount of theactive agent or agents without undue experimentation.

EXAMPLES

Having now described the several embodiments of the present disclosurein detail, the same will be more clearly understood by reference to thefollowing examples, which are included herewith for purposes ofillustration only and are not intended to be limiting of the presentdisclosure.

Example 1. Synthesis of Telmisartan/Tempol Conjugate (YK-4-250)

Analytical Methods. NMR spectra were recorded using a Varian-400spectrometer for ¹H (400 MHz). Chemical shifts (δ) are given in ppmdownfield from tetramethylsilane, as internal standard, and couplingconstants (J-values) are in hertz (Hz). Purifications by flashchromatography were performed. Liquid chromatography/mass spectrometry(LC/MS) analyses were conducted using Shimadzu LC-20AD pumps and aSPD-20A UV-vis detector. High-resolution mass spectra (HMRS) wererecorded on a QSTAR Elite mass spectrometer.

Telmisartan Extraction. Telmisartan tablets were triturated, suspendedin methanol and stirred for about 20 mins. Filtered off the solid, themethanol solution was concentrated, and the residue was purified bychromatography to afford white solid in 90% yield. ¹H NMR (CDCl₃, 400MHz) δ 8.38 (m, 1H), 8.02 (dd, 1H, J=1.2, 1.2 Hz), 7.39 (m, 8H), 7.17(s, 1H), 7.15 (s, 1H), 7.04 (s, 1H), 6.95 (s, 1H), 5.40 (s, 2H), 3.74(s, 3H), 3.13 (t, 2H, J=7.6, 8.0 Hz), 2.69 (s, 3H), 1.99 (m, 2H), 1.15(t, 3H, J=7.6, 7.2 Hz).

Conjugation of Telmisartan with Tempol (YK-4-250).

To an ice bath cooled solution of telmisartan (0.8 g, 1.55 mmol) in DMF(50 mL) was added 1-hydroxybenzotriazole (HOBt, 0.25 g, 1.87 mmol),4-dimethylamino pyridine (DMAP, 0.23 g, 1.87 mmol), and1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDCI, 0.39g, 2.02 mmol), followed by tempol (0.29 g, 1.712 mmol). The mixture wasstirred at room temperature for 48 h. Water (15 mL) was added to themixture and stirred at room temperature for 10 minutes. The mixture wasextracted with ethyl acetate (3×15 mL). The organic layer was washedwith sat. LiCl (15 mL), sat. NaHCO₃ (15 mL), water (15 mL) and brine (15mL), dried over Na2SO₄. The solvent was evaporated and the residue waspurified by flash chromatography using CH₂Cl₂-MeOH to afford YK-4-250 asa pink soft solid (0.81 g, 78%). LC-MS (ESI): m/z 669 (M+H)⁺; HRMS(TOF): calculated for C₄₂H₄₇N₅O₃ (M+H)⁺: 669.3679; Found: 669.3578.

Example 2. In Vitro Protective Effects of Telmisartan

Gamma radiation: YAMC (normal colonic epithelial cell line) were exposedto 4 Gy gamma ionizing radiation (IR) with air pumped into the chamberduring exposure. A Gammacell 40 ¹³⁷Cs gamma irradiator was used with adose rate of 0.8 Gy IR per minute. Dosimetry measurements were performedusing Fricke Dosimetry systems. Measured absorbance dose was: CentralDose Rate (0.790 Gy/min±2.9%); 3 o'clock position (0.804 Gy/min±3.8%);and 7 o'clock position (0.808 Gy/min±2.7%). The experiments wereperformed within 6 months of the dosimetry analysis. YAMC cells in thetreatment group received telmisartan 24 h post-IR at three differentdoses (10 μM, 100 μM, and 1 mM).

Colony forming assay: Colony forming assay is universally recognized asthe gold standard method for measuring the effects of radiation,chemotherapeutic drugs on cell viability, and cell survival. 24 hpost-IR YAMC cells were split into 6-well plates for colony formationassessment with and without telmisartan treatment. At the end of 96 h,cells were fixed and counted for number of colonies formed. The numberof colonies formed represents the number of cells survived.Representative results are shown in FIG. 1 . As shown in the figure,telmisartan treatment significantly increased YAMC cell survivalcompared to vehicle treatment after 4 Gy IR.

Clonogenic assay for self-renewal: Self-renewal is the hallmark for stemcells or stem-like cells. Clonogenic assay or clonal expansion of singlecell into spheroids/organoids is the reputed method to analyze cellself-renewal. 24 h post-IR YAMC cells were split into 96-well plates(ultra-low attachment plates) in spheroid growth conditioned media forself-renewal ability with and without telmisartan. At the end of 192 h,spheroids were counted for quantitative assessment of the number ofspheroids formed. Representative results are shown in FIG. 2 . As shownin the figure, telmisartan treatment significantly increased theself-renewal ability of YAMC cells compared to vehicle treatment after 4Gy IR.

Example 3. In Vivo Protective Effects of Telmisartan

Gamma radiation: 7-week-old C57BL/6 mice were exposed to total bodyirradiation (TBI) 14 Gy gamma ionizing radiation (IR) with air pumpedinto the chamber during. A Gammacell 40 ¹³⁷Cs gamma irradiator was usedwith a dose rate of 0.8 Gy IR per minute. Dosimetry measurements wereperformed using Fricke Dosimetry systems. Measured absorbance dose was:Central Dose Rate (0.790 Gy/min±2.9%); 3 o'clock position (0.804Gy/min±3.8%); and 7 o'clock position (0.808 Gy/min±2.7%). Theexperiments were performed within 6 months of the dosimetry analysis.All IR treatments were begun in the morning. Animals in the treatmentgroup received three doses of telmisartan, 10 mg/kg body weight per day,beginning 24 h post-IR. All animals received easy access to liquid gelfood and water in the cage.

Crypt survival analysis: Two hours before euthanasia (82 h post-IR),each mouse was intraperitoneally injected with 5-bromo-2′-deoxyuridine(BrdUrd, Sigma Aldrich, St. Louis, MO; 200 μL of 5 mg/mL BrdUrd solutionin PBS).

Barrier function study: Two hours before euthanasia (5 d post-IR), eachmouse was orally administered (gavage) with 100 μL of FITC-dextran (FITCdextran 4; MW 4000) at a dose of 80 mg/100 g body weight. Serum analysisof FITC concentration was performed in triplicate using Synergy Bio TEKplate reader. Representative results are shown in FIG. 3 . As shown inthe figure, intestinal epithelial permeability to FITC-dextran wasessentially the same for vehicle control and telmisartan groups.

Overall survival study: Animals following treatment (IR and/orIR+telmisartan) were allowed to survive. Animals demonstrating bloodystools, lethargy, or unable to move were killed and time of death noted.Survival time of mice in the treatment group was compared with controlIR-alone-treated mice. Representative results are shown in FIG. 4 . Asshown in the figure, median survival for control-treated mice was 204days, whereas median survival for telmisartan-treated mice was 264 days.Log-rank (Mantel-Cox) test chi square 7.962; p=0.0048.

Example 4. In Vitro Protective Effects of Tempol

Gamma radiation: YAMC (normal colonic epithelial cell line) were exposedto 4 Gy gamma ionizing radiation (IR) with air pumped into the chamberduring exposure. A Gammacell 40 ¹³⁷Cs gamma irradiator was used with adose rate of 0.8 Gy IR per minute. Dosimetry measurements were performedusing Fricke Dosimetry systems. Measured absorbance dose was: CentralDose Rate (0.790 Gy/min±2.9%); 3 o'clock position (0.804 Gy/min±3.8%);and 7 o'clock position (0.808 Gy/min±2.7%). The experiments wereperformed within 6 months of the dosimetry analysis. YAMC cells in thetreatment group received tempol 24 h post-IR at three different doses(10 μM, 100 μM, and 1 mM).

Colony forming assay: Colony forming assay is universally recognized asthe gold standard method for measuring the effects of radiation,chemotherapeutic drugs on cell viability, and cell survival. 24 hpost-IR YAMC cells were split into 6-well plates for colony formationassessment with and without tempol treatment. At the end of 96 h, cellswere fixed and counted for number of colonies formed. The number ofcolonies formed represents the number of cells survived. Representativeresults are shown in FIG. 5 . As shown in the figure, tempol treatmentsignificantly increased YAMC cell survival compared to vehicle treatmentafter 4 Gy IR.

Clonogenic assay for self-renewal: Self-renewal is the hallmark for stemcells or stem-like cells. Clonogenic assay or clonal expansion of singlecell into spheroids/organoids is the reputed method to analyze cellself-renewal. 24 h post-IR YAMC cells were split into 96-well plates(ultra-low attachment plates) in spheroid growth conditioned media forself-renewal ability with and without tempol. At the end of 192 h,spheroids were counted for quantitative assessment of the number ofspheroids formed. Representative results are shown in FIG. 6 . As shownin the figure, tempol treatment significantly increased the self-renewalability of YAMC cells compared to vehicle treatment after 4 Gy IR.

Example 5. In Vivo Protective Effects of Tempol

Gamma radiation: 7-week-old C57BL/6 mice were exposed to total bodyirradiation (TBI) 14 Gy gamma ionizing radiation (IR) with air pumpedinto the chamber during. A Gammacell 40 ¹³⁷Cs gamma irradiator was usedwith a dose rate of 0.8 Gy IR per minute. Dosimetry measurements wereperformed using Fricke Dosimetry systems. Measured absorbance dose was:Central Dose Rate (0.790 Gy/min±2.9%); 3 o'clock position (0.804Gy/min±3.8%); and 7 o'clock position (0.808 Gy/min±2.7%). Theexperiments were performed within 6 months of the dosimetry analysis.All IR treatments were begun in the morning. Animals in the treatmentgroup received three doses of tempol, 10 mg/kg body weight per day,beginning 24 h post-IR. All animals received easy access to liquid gelfood and water in the cage.

Crypt survival analysis: Two hours before euthanasia (82 h post-IR),each mouse was intraperitoneally injected with 5-bromo-2′-deoxyuridine(BrdUrd, Sigma Aldrich, St. Louis, MO; 200 μL of 5 mg/mL BrdUrd solutionin PBS).

Barrier function study: Two hours before euthanasia (5 d post-IR), eachmouse was orally administered (gavage) with 100 μL of FITC-dextran (FITCdextran 4; MW 4000) at a dose of 80 mg/100 g body weight. Serum analysisof FITC concentration was performed in triplicate using Synergy Bio TEKplate reader. Representative results are shown in FIG. 7 . As shown inthe figure, intestinal epithelial permeability to FITC-dextran wasessentially the same for vehicle control and tempol groups.

Overall survival study: Animals following treatment (IR and/orIR+tempol) were allowed to survive. Animals demonstrating bloody stools,lethargy, or unable to move were killed and time of death noted.Survival time of mice in the treatment group was compared with controlIR-alone-treated mice. Representative results are shown in FIG. 8 . Asshown in the figure, median survival for control-treated mice was 204days, whereas median survival for tempol-treated mice was 276 days.Log-rank (Mantel-Cox) test chi square 7.962; p=0.0048.

Example 6. In Vitro Protective Effects of YK-4-250

Gamma radiation: YAMC (normal colonic epithelial cell line) were exposedto 4 Gy gamma ionizing radiation (IR) with air pumped into the chamberduring exposure. A Gammacell 40 ¹³⁷Cs gamma irradiator was used with adose rate of 0.8 Gy IR per minute. Dosimetry measurements were performedusing Fricke Dosimetry systems. Measured absorbance dose was: CentralDose Rate (0.790 Gy/min±2.9%); 3 o'clock position (0.804 Gy/min±3.8%);and 7 o'clock position (0.808 Gy/min±2.7%). The experiments wereperformed within 6 months of the dosimetry analysis. YAMC cells in thetreatment group received YK-4-250 24 h post-IR at three different doses(10 μM, 100 μM, and 1 mM).

Colony forming assay: Colony forming assay is universally recognized asthe gold standard method for measuring the effects of radiation,chemotherapeutic drugs on cell viability, and cell survival. 24 hpost-IR YAMC cells were split into 6-well plates for colony formationassessment with and without YK-4-250 treatment. At the end of 96 h,cells were fixed and counted for number of colonies formed. The numberof colonies formed represents the number of cells survived.Representative results are shown in FIG. 9 . As shown in the figure,YK-4-250 treatment significantly increased YAMC cell survival comparedto vehicle treatment after 4 Gy IR.

Clonogenic assay for self-renewal: Self-renewal is the hallmark for stemcells or stem-like cells. Clonogenic assay or clonal expansion of singlecell into spheroids/organoids is the reputed method to analyze cellself-renewal. 24 h post-IR YAMC cells were split into 96-well plates(ultra-low attachment plates) in spheroid growth conditioned media forself-renewal ability with and without YK-4-250. At the end of 192 h,spheroids were counted for quantitative assessment of the number ofspheroids formed. Representative results are shown in FIG. 10 . As shownin the figure, YK-4-250 treatment significantly increased theself-renewal ability of YAMC cells compared to vehicle treatment after 4Gy IR.

Example 7. In Vivo Protective Effects of YK-4-250

Gamma radiation: 7-week-old C57BL/6 mice were exposed to total bodyirradiation (TBI) 14 Gy gamma ionizing radiation (IR) with air pumpedinto the chamber during. A Gammacell 40 ¹³⁷Cs gamma irradiator was usedwith a dose rate of 0.8 Gy IR per minute. Dosimetry measurements wereperformed using Fricke Dosimetry systems. Measured absorbance dose was:Central Dose Rate (0.790 Gy/min±2.9%); 3 o'clock position (0.804Gy/min±3.8%); and 7 o'clock position (0.808 Gy/min±2.7%). Theexperiments were performed within 6 months of the dosimetry analysis.All IR treatments were begun in the morning. Animals in the treatmentgroup received three doses of YK-4-250, 10 mg/kg body weight per day,beginning 24 h post-IR. All animals received easy access to liquid gelfood and water in the cage.

Crypt survival analysis: Two hours before euthanasia (82 h post-IR),each mouse was intraperitoneally injected with 5-bromo-2′-deoxyuridine(BrdUrd, Sigma Aldrich, St. Louis, MO; 200 μL of 5 mg/mL BrdUrd solutionin PBS).

Barrier function study: Two hours before euthanasia (5 d post-IR), eachmouse was orally administered (gavage) with 100 μL of FITC-dextran (FITCdextran 4; MW 4000) at a dose of 80 mg/100 g body weight. Serum analysisof FITC concentration was performed in triplicate using Synergy Bio TEKplate reader. Representative results are shown in FIG. 11 . As shown inthe figure, intestinal epithelial permeability to FITC-dextran wasessentially the same for vehicle control and YK-4-250 groups.

Overall survival study: Animals following treatment (IR and/orIR+YK-4-250) were allowed to survive. Animals demonstrating bloodystools, lethargy, or unable to move were killed and time of death noted.Survival time of mice in the treatment group was compared with controlIR-alone-treated mice. Representative results are shown in FIG. 12 . Asshown in the figure, median survival for control-treated mice was 204days, whereas median survival for YK-4-250-treated mice was 240 days.Log-rank (Mantel-Cox) test chi square 3.891; p=0.0486.

INCORPORATION BY REFERENCE

All patents and published patent applications mentioned in thedescription above are incorporated by reference herein in theirentireties.

EQUIVALENTS

Having now fully described several embodiments of the present disclosurein some detail by way of illustration and example for purposes ofclarity of understanding, it will be obvious to one of ordinary skill inthe art that the same can be performed by modifying or changing thevarious embodiments within a wide and equivalent range of conditions,formulations and other parameters without affecting the scope of thedisclosure or any specific embodiment thereof, and that suchmodifications or changes are intended to be encompassed within the scopeof the appended claims.

1.-12. (canceled)
 13. A method for treating or preventingradiation-induced tissue injury, comprising administering to a subjectin need thereof a therapeutically effective amount of a compoundrepresented by formula II

or a pharmaceutically acceptable salt thereof, thereby treating orpreventing radiation-induced tissue injury in the subject.
 14. Themethod of claim 13, wherein the compound is administered prior toexposure of the subject to radiation.
 15. The method of claim 13,wherein the compound is administered after exposure of the subject toradiation.
 16. The method of claim 13, wherein the radiation-inducedtissue injury is acute radiation-induced tissue injury.
 17. The methodof claim 13, wherein the radiation-induced tissue injury is chronicradiation-induced tissue injury.
 18. The method of claim 13, wherein thetissue comprises gastrointestinal tissue.
 19. The method of claim 13,wherein the tissue comprises hematopoietic tissue.
 20. The method ofclaim 13, wherein the tissue comprises neural tissue.
 21. The method ofclaim 13, wherein the compound is administered orally.
 22. The method ofclaim 13, wherein the compound is administered parenterally. 23.-42.(canceled)
 43. The method of claim 13, wherein the subject is a mammal.44. The method of claim 13, wherein the subject is a human.
 45. Themethod of claim 13, wherein the radiation-induced tissue injury isprevented.
 46. The method of claim 13, wherein the radiation-inducedtissue injury is treated.
 47. The method of claim 14, wherein theradiation-induced tissue injury is acute radiation-induced tissueinjury.
 48. The method of claim 14, wherein the radiation-induced tissueinjury is chronic radiation-induced tissue injury.
 49. The method ofclaim 15, wherein the radiation-induced tissue injury is acuteradiation-induced tissue injury.
 50. The method of claim 15, wherein theradiation-induced tissue injury is chronic radiation-induced tissueinjury.